Cellular Mechanisms of PTLD in Transplant Recipients

描述(申请人提供)：移植后淋巴增生性疾病(PTLD)描述了实体器官和骨髓移植的形态异质性并发症，范围从良性B细胞增生到恶性淋巴瘤。PTLD的特点是发生与EB病毒相关的B细胞淋巴瘤。目前，PTLD还没有成熟的治疗方法，报道的死亡率通常超过50%。治疗PTLD的策略有多种，如抗病毒药物、B细胞特异性抗体，包括抗CD20(利妥昔单抗)和过继细胞免疫治疗。然而，这些方法都有明显的局限性。我们广泛的、长期的目标是确定对PTLD相关的EBV+B细胞淋巴瘤的生长和生存至关重要的细胞和病毒蛋白，以期确定治疗靶向的新候选者。我们假设EBV通过宿主细胞途径促进B细胞肿瘤的生长和存活。为了支持这一假设，我们定义了一条新的途径，即潜伏期EBV蛋白，潜伏膜蛋白1(LMP1)，通过它在感染的B细胞中诱导产生人IL-10(hIL-10)。IL-10在包括PTLD在内的多种EBV+相关的恶性肿瘤中发挥着强大的自分泌生长因子的作用。我们还确定了由病毒蛋白潜伏膜蛋白2A(LMP2A)激活的脾酪氨酸激酶(Syk)对PTLD相关B细胞系的生存至关重要。此外，我们的初步数据表明，Syk的小分子抑制剂显著抑制PTLD来源的细胞系的生长和存活。最后，我们证明了LMP1通过对CTL和NK细胞用来清除病毒感染和转化细胞的关键死亡受体(DR)效应机制的抵抗，通过依赖于NF：B的途径来颠覆宿主免疫。我们的发现表明，细胞蛋白cFLIP参与了抵抗DR诱导的细胞凋亡。在这项提案中，我们将针对已识别的信号转导途径和细胞蛋白，由关键病毒蛋白LMP1和LMP2a诱导，目的是抑制PTLD相关B细胞淋巴瘤的生长。为了实现这些目标，我们提出了三个具体目标。具体目标1将研究Syk在PTLD相关EBV+B细胞淋巴瘤生存中的需求，并在NOD/SCID模型中测试以Syk为靶点作为潜在治疗策略的价值。特异性目标2将确定LMP1介导的PI3K/AKT信号轴的激活及其在hIL-10诱导中的作用。此外，干预这一途径的策略将作为一种抑制体内肿瘤生长的手段进行测试。在特定的目标3中，我们将靶向LMP1诱导的细胞蛋白cFLIP，目的是提高宿主CTL消除EBV感染的B细胞淋巴瘤的能力。我们预计，这些方法将为PTLD的治疗提供新的、有效的治疗选择。 公共卫生相关性：爱泼斯坦-巴尔病毒相关的B细胞淋巴瘤的发展仍然是实体器官和骨髓移植的严重并发症，而且往往是致命的。目前，还没有治疗这种疾病的既定疗法。因此，识别和定义抑制肿瘤生长的新靶点的研究可能会导致新的诊断和治疗药物来预防这种毁灭性的疾病。

DESCRIPTION (provided by applicant): Post-transplant lymphoproliferative disease (PTLD) describes a morphologically heterogeneous complication of solid organ and bone marrow transplantation that ranges from benign B cell hyperplasia to malignant lymphoma. The hallmark of PTLD is the development of Epstein Barr Virus (EBV)-associated B cell lymphomas. Currently, there are no established therapies for PTLD and reported mortality rates commonly exceed 50%. Multiple strategies have been utilized to treat patients with PTLD such as anti-viral drugs, B cell specific antibodies including anti-CD20 (Rituximab) and adoptive cellular immunotherapy. However, each of these approaches has distinct limitations. Our broad, long term objective has been to determine the cellular and viral proteins that are essential to the growth and survival of PTLD-associated EBV+ B cell lymphomas with the intent to identify novel candidates for therapeutic targeting. We hypothesize that EBV co-opts host cellular pathways to promote growth and survival of B cell tumors. In support of this hypothesis, we have defined a novel pathway by which the latent cycle EBV protein, latent membrane protein 1 (LMP1) induces production of human IL-10 (hIL-10) in infected B cells. hIL-10 acts as a potent autocrine growth factor in a variety of EBV+- associated malignancies including PTLD. We have also determined that the spleen tyrosine kinase (Syk), activated by the viral protein latent membrane protein 2A (LMP2A), is critical to the survival of PTLD- associated B cell lines. Moreover, our preliminary data indicate that a small molecule inhibitor of Syk significantly inhibits the growth and survival of PTLD-derived cell lines. Finally we demonstrate that LMP1 subverts host immunity through an NF:B-dependent pathway by conferring resistance to key death-receptor (DR) effector mechanisms utilized by CTL and NK cell to eliminate virally-infected and transformed cells. Our findings implicate the cellular protein, cFLIP in the resistance to DR-induced apoptosis. In this proposal we will target identified signal transduction pathways, and cellular proteins, induced by the key viral proteins LMP1 and LMP2a with the goal of inhibiting growth of PTLD-associated B cell lymphomas. To achieve these objectives we propose three specific aims. Specific Aim 1 will investigate the requirement for Syk in survival of PTLD-associated EBV+ B cell lymphomas and test, in a NOD/SCID model, the value of targeting Syk as a potential therapeutic strategy. Specific Aim 2 will define the LMP1-mediated activation of the PI3K/AKT signaling axis and its role in hIL-10 induction. Further, strategies to intervene in this pathway will be tested as a means to inhibit tumor growth in vivo. In Specific Aim 3, we will target the LMP1-induced cellular protein, cFLIP with the intent to boost the ability of host CTL to eliminate EBV-infected B cell lymphomas. We anticipate that these approaches will reveal novel, and effective, therapeutic options for the treatment of PTLD.7. PUBLIC HEALTH RELEVANCE: The development of Epstein Barr Virus-associated B cell lymphomas remains a serious, and often fatal, complication of solid organ and bone marrow transplantation. Currently, there are no established therapies for treatment of this disease. Thus, studies that identify and define novel targets for inhibiting tumor growth could lead to new diagnostic and therapeutic agents for prevention of this devastating disease.